This proposal explores the cellular and molecular mechanisms of a non-tissue damaging continuous wave (CW) near-infrared (NIR) laser to enhance adaptive immune responses, and develops a safe and effective immunologic adjuvant for skin and mucosal vaccination. The ultimate purpose of this is to replace a concurrent chemical or biological vaccine adjuvant. Although immunologic adjuvants are often essential for current vaccines, many licensed and development-stage adjuvants cause significant side effects. This necessitates the development of new safe and effective adjuvants. Illumination of the skin with nanosecond-pulsed wave (PW) visible range laser light has previously been shown to enhance immune responses to vaccination in humans and mice. However, these lasers require co-administration of an additional chemical adjuvant to induce a sufficiently increased immunological response. In addition, laser light in the green or yellow spectrums is absorbed by melanin, resulting in highly variable light absorption across different skin phototypes and therefore limiting its clinical utility. We have shown, for the first time, that that non-tissue damaging NIR laser light given in short exposures to small areas of the skin, without the use of any additional agents, increases a broad spectrum of immune responses to influenza antigen. This occurs at a magnitude comparable to licensed adjuvants, and results in improved survival in a lethal challenge murine model. Use of a low-power CW NIR laser has several advantages over PW visible light lasers, as well as licensed chemical vaccine adjuvants. Since water is the predominant chromophore for NIR laser light, light absorption is not significantly altered across different skin phototypes. The laser reduces potential adjuvant reactogenicity and toxicity, as it does not induce a prolonged inflammatory cytokine response, does not promote allergenicity, and unlike chemical adjuvants, does not persist in exposed tissues. Since the laser is external to a vaccine it does not pose any stability issues that conventional vaccine-chemical adjuvant combinations do. These provocative findings raise the following three important areas for further investigation that we will address in this proposal: 1) To explore whether NIR laser light in the skin enhances a long-term recall immune response to skin-based influenza vaccination using established animal models of influenza infection in mice and ferrets, 2) Elucidation of the cellula and molecular mechanisms of action of the NIR laser adjuvant in vitro and in vivo, and 3) Exploration of the use of this technology in mucosal vaccinations in mice and ferrets. Upon the culmination of this proposal, we aim to demonstrate the efficacy and mechanisms of action of the NIR laser adjuvant in the context of skin and mucosal vaccination. The translatability of this discovery is enhanced by the current availability of mature, safe, compact and relatively simple low-wattage CW NIR laser technology, making it possible to economically produce a portable (handheld) low cost device that, at sufficient vaccination volumes, could replace chemical adjuvants. Such a device could have an immediate and tremendous impact on current clinical practice.